BCR-ABL as a Molecular Target
Jorge Cortes, Michael Deininger in Chronic Myeloid Leukemia, 2006
Abl (also referred to as Abl-1), the human homologue of the Abelson murine leukemia virus, is a tyrosine kinase involved in multiple cellular processes, including DNA repair, integrin signaling, cell cycle regulation, and signal transduction from cell surface receptors (11). ABL knockout mice exhibit increased neonatal lethality and suffer from a number of defects, including skeletal malformations, immune dysfunction as well as an ill-defined wasting syndrome (12,13). Apart from the unique “Cap” region at its very 5’ end the N-terminus of Abl has extensive homology to Src kinases (14). The Src homology domains 3 (SH3) and 2 (SH2) mediate interactions with other proteins by binding proline rich regions (SH3) or phosphotyrosine (SH2). The SH1 domain carries the tyrosine kinase function. The large C-terminus is unique to Abl and contains DNA binding, nuclear localization, and export signals as well as actin-binding sequences and a proline rich domain(15). In physiological conditions, Abl kinase is tightly regulated by a mechanism that is similar as in Src kinases but uses different structures, as Abl lacks a C-terminal tyrosine that is critical for auto-inhibition of Src kinases (14). In Src, phosphorylation of tyrosine 527 allows the N-terminus to form an intramolecular association with the SH2 domain, inactivating the kinase by forcing the molecule into a “clamp.” In Abl, the myristoylated cap binds to a hydrophobic pocket at the base of the kinase domain, resulting in a conformation resembling inactive SRC.
Treatment of Chronic Myeloid Leukemia with Bcr-Abl Kinase Inhibitors
Gertjan J. L. Kaspers, Bertrand Coiffier, Michael C. Heinrich, Elihu Estey in Innovative Leukemia and Lymphoma Therapy, 2019
Several important experimental works established the ability of Bcr-Abl, as a singular oncogenic abnormality, to cause leukemia. Transgenic mice that express the p190 Bcr-Abl fusion protein were shown to develop a rapidly fatal acute leukemia (13). Transduction of p210 Bcr-Abl into murine hematopoetic stem cells, followed by transplantation into syngeneic mice, causes a CML-like syndrome (14,15). Although convincing, there was and remains controversy about the genesis of CML, particularly surrounding the possibility of clonal stem cell events antecedent to 9:22 translocation and Bcr-Abl fusion, which was championed by Fiaklow and others decades ago (16). More recent work has bolstered the transforming potential of Bcr-Abl by demonstrating that mice expressing a Bcr-Abl transgene, under the control of a tetracycline-repressible promoter, develop a reversible leukemia entirely dependent on the presence or absence of tetracycline (17).
Oncogenes and Cancer
Pimentel Enrique in Oncogenes, 2020
In another study fresh human leukemia cells from different types of leukemia were obtained from leukapheresis in a total of 26 patients and human c-onc probes, not v-onc probes, were used for determining the expression of different proto-oncogenes by molecular hybridization.70 It was found that “different oncogenes are expressed in the different leukemic types and that the transcript copy number varies from one type of leukemia to another and within a given individual leukemic type”.70 The c-myc gene was expressed at detectable levels in all leukemia samples analyzed and the c-myb gene was expressed in all samples except in B-cell malignancies. A single 1.5 kb transcript of c-H-ras was present at low levels in all acute and chronic leukemias examined whereas c-sis was expressed only in the sample from one patient with chronic myeloid leukemia in blast transformation. This is the first reported case of c-sis transcriptional activity in human malignant diseases. The proto-oncogene c-abl was expressed at very low levels in a fraction of all leukemia types analyzed and the transcript sizes and levels were the same in chronic myeloid leukemia (where the c-abl gene is translocated) as in other forms of leukemia. However, in some samples of CML an aberrant 8.0 kb transcript was detected in addition to the normal 6.4 and 7.2 kb transcripts contained in control samples. The scr and erb proto-oncogenes were not expressed in the fresh human leukemia cell samples.
Antiproliferative Effect of Gaillardin from Inula oculus-christi in Human Leukemic Cells
Published in Nutrition and Cancer, 2020
Afshin Karami, Maryam Hamzeloo-Moghadam, Amir Yami, Mohieddin Barzegar, Pargol Mashati, Ahmad Gharehbaghian
Initially, cells (3 × 105/mL) were treated with various concentrations of VCR and GLN, and were incubated for 48 h. Total RNA was extracted from each well by the Hybrid-R RNA purification kit (Gene All, Korea) according to the manufacturer’s instructions. The quantity of the RNA samples was analyzed by UV-spectroscopy (Nano Drop TM 2000 Spectrophotometer, Thermo Scientific, USA) at 260 and 280 nm and the quality of extracted RNA was tested by the 1% agarose gel electrophoresis. RNA was reverse transcribed to first-strand cDNA using the Revert Aid First Strand cDNA Synthesis Kit (Thermo Scientific, USA) following the manufacturer’s instructions. The sequences of the primers are shown in Table 1. Real-time PCR analysis was performed using SYBR Premix Ex Taq (Tli RNase H Plus) kit (Takara Biomedical Technology, Japan) in Applied Rotor-Gene Q Real-Time PCR System (Qiagen, Valencia, CA). The expression of ABL gene was used as an endogenous control to normalize the expression of the target genes. Melting curve analysis of CASPASE-3, BCL-2, BAX, P21, C-MYC, and ABL showed a single peak. The mean Ct of the target genes was calculated from triplicate measurements and then normalized with the mean Ct of the ABL gene. Data extraction was executed using the Rotor-Gene Q series software v. 2.0.2, and the Livak method was used to calculate the relative expression of each gene (20).
Insights into apoptotic proteins in chemotherapy: quantification techniques and informing therapy choice
Published in Expert Review of Proteomics, 2018
BCR-ABL mutations belong to the most characteristic genetic alterations in chronic myeloid leukemia (CML). BCR-ABL inhibition can be achieved by combination treatment with several tyrosine kinase inhibitors to exploit their synergistic potential. A synergy was found between danusertib (an aurora kinase inhibitor) and bosutinib (a multiple tyrosine kinase inhibitor) that affected CML cells characterized by BCR-ABL mutations. Phosphoproteomics, transcriptomics and chemical proteomics were used to detect the underlying mechanisms. Both compounds targeted MAPK pathways downstream of BCR-ABL, resulting in impaired activity of c-Myc. Using pharmacological validation, we assessed that the relative contributions of danusertib and bosutinib could be individually mimicked by MAPK inhibitors and collectively by downregulation of c-Myc through Brd4 inhibition [76].
Anticancer Activity and Molecular Mechanism of Momordica cochinchinensis Seed Extract in Chronic Myeloid Leukemia Cells
Published in Nutrition and Cancer, 2022
Zhengdong Ai, Chong Ma, Ruiming Wan, Jingyi Yin, Guiming Li, Yan Li, Li Chen
The formation of Bcr-Abl is the pathogenesis of chronic myeloid leukemia. Targeting Bcr-Abl is the key to treating CML patients. Therefore, a plasmid for determining the reporter activity of the Bcr-Abl promoter was established by incorporating the Bcr-Abl promoter region (approximately 2000 base pairs) into firefly luciferase expression plasmid (pGL3-Basic, Promega), by which the inhibitory activity of MCSE against Bcr-Abl transcription was examined. The results demonstrated that MCSE significantly inhibited the Bcr-Abl promoter activity (Figure 2A) in HEK-293T cells. Meanwhile, the expression levels of Bcr-Abl in two CML cell lines KBM5 and KBM5-T315I were investigated after CML cells were treated with different concentrations (0, 20, 40 and 60 μg/ml) of MCSE by RT-qPCR. The results were consistent with expectations that MCSE effectively inhibited the transcription of the Bcr-Abl gene in a concentration-dependent manner (Figure 2B). The inhibitory effect of MCSE on Bcr-Abl transcription in CML cells is stronger than that of MCSE on Bcr-Abl promoter activity in HEK-293T cells, indicating that MCSE may affect the level of Bcr-Abl mRNA from other aspects such as stability in CML cells.
Related Knowledge Centers
- Abelson Murine Leukemia Virus
- Cell Adhesion
- Cell Division
- Chromosome 9
- DNA Repair
- Oncogene
- Protein
- Sh3 Domain
- Gene
- Tyrosine Kinase